Many human genetic diseases are due to the deficiency of an enzyme or other protein. For example, the genetically determined deficiency of the lysosomal enzyme alpha-L-iduronidase results in the progressive accumulation of glycosaminoglycan substrates. In vitro evidence that cells grown in culture can take up exogenously supplied enzymes from the surrounding tissue culture medium led to the concept of in vivo enzyme replacement. For instance, enzyme replacement by intravenous infusion has been demonstrated to be successful for adenosine deaminase deficiency and for Gaucher disease, and some measures of efficacy have been found in human patients receiving weekly infusions of recombinant human alpha-L-iduronidase. However, these infusions must occur over a period of hours every week, and it is unclear if weekly administration of the enzyme would be efficacious, even if treatments are started early in life. Moreover, it is likely that enzyme infusions will not prevent progressive mental retardation associated with particular protein deficiencies.
Enzyme replacement may also be accomplished by transplantation of genetically normal cells and tissues, e.g., via bone marrow transplantation for mucopolysaccharidosis (also known as Hurler syndrome). For example, bone marrow transplantation was found to reduce many of the consequences of mucopolysaccharidosis type I and may prevent progression of mental retardation (Whitley et al., 1986). However, transplantation procedures which include the use of immunosuppressive medications are associated with an increase in morbidity and mortality.
Enzyme replacement may also be accomplished via gene therapy e.g., with viral vectors such as HIV-based vectors, ex vivo or in vivo. Lentiviral vectors are one type of viral vector which has been proposed as useful for mammalian gene therapy. HIV-based lentiviral vectors pseudotyped with the envelope of another virus (most often the G protein of the vesicular stomatitis virus, VSVG) have become promising tools for gene delivery into nondividing cells. These vectors have been shown to be capable of transferring genes into a range of nonproliferative cell types, including neurons, retinal cells, muscle cells, and hematopoietic pluripotent cells (Amado et al., 1999; Lever, 2000; Podsakoff, 2001) and, using local administration of those vectors, in vivo gene delivery has been accomplished in rat brain (Naldini et al., 1996; Blomer et al., 1997), liver and muscle (Kafri et al., 1997), retina (Miyoshi et al., 1999), and airway epithelia (Johnson et al., 2000).
However, the biosafety concerns surrounding HIV vectors have received considerable attention because of the pathogenic nature of HIV. Thus, efforts have been made to increase the safety of lentivirus vectors by minimizing the potential formation of replication-competent virus (RCR) via homologous recombination events. One strategy to reduce RCR formation has been to use nonoverlapping split-genome packaging constructs that require multiple recombination events with the transfer vector for RCR generation (Naldini et al., 1996; Reiser et al., 1996). Other strategies have focused on eliminating all unnecessary HIV reading frames from the system (Kim et al., 1998; Dull et al., 1998) or truncating the 3′ long terminal repeat (3′ LTR) to generate self-inactivating HIV vectors (Miyoshi et al., 1998).
Lentiviral vectors have been a preferred vector for ex vivo modification of hematopoietic (i.e., blood-forming) stem cells as lentiviral vectors are likely capable of transducing such nondividing types of cells. Nevertheless, despite thousands of experiments attempting ex vivo gene transfer into hematopoietic stem cells, this vector-cell combination has not been successful in animal models of disease. Moreover, there has never been a successful clinical response in an animal using in vivo lentiviral gene therapy, probably owing to insufficient delivery of vector, or lack of expression of therapeutic protein, to the disease-causing tissues or cells.
Thus, what is needed is an improved method to prevent, inhibit or treat metabolic disorders characterized by a lack of, or a reduction in the amount of, an enzyme in a mammal.